Time-Resolved Fluorescence Spectroscopy is a powerful tool for biochemistry;it can provide unique insights into the structure and assembly of macromolecular complexes. This year, we pursued protein-protein association , ultrafast protein solvation, and probes of mitochondrial energetics. We also continued studies of DNA using fluorescent nucleotide analogs that reveal disruptions in DNA structures, publishing accounts of analog base pairing effects. We continued and expanded our femtosecond upconversion studies of Trp in proteins and peptides to quantify early "quasistatic self-quenching" processes. We found extremely rapid (10-100ps) decays are important in several proteins (crystallins, thioredoxin, GB1,etc.), as they detect previously silent conformers engaged in ultrafast charge transfer. Our earlier study of protein *solvation* on the 330fs-200ps time scale, using proteins such as Monellin, found QSSQ that others attributed to a class of unique water molecules that desorb from protein in 20ps. Local quenching is the dominant mechanism in all but a few cases we have studied. The QSSQ was found even in simple dipeptides (just published)(suggesting a general process underlies all protein QSSQ) and we suggest that in crystallins QSSQ is an important factor in preventing eye lens cataract formation. We contined collaborative studies with LCE into the status of a primary fuel of heart muscle mitochondria- NADH. Our efforts distinguish free and bound populations of NADH by their different fluorescence lifetimes, and in collaboration with Light Microscopy Core and LCE, we are refining 'Decay-Associated Images'software to more rapidly extract profiles of NADH binding within isolated cardiac myocytes. We studied the structural determinants of lifetime for pteridine-based fluorophores that mimic (to the point of base-pairing) the behavior of natural DNA bases, learning which neighbors provide a rigid base "sandwich" and which permit "flip-out". We employed the same probes to look at the G-quadruplex formation within DNA (important in teleomeres , division, and regulation).